It has been well known that adult stem cells promise to have tremendous therapeutic potential in regenerative medicine due to their plasticity of differentiation into multiple cell lineages. In bone marrow, there are certain populations of stem cell sources including hematopoietic stem cells, mesenchymal stem cells (MSCs), and multi-potent adult progenitor cells [1. Evans M, Kaufman M. Establishment in culture of pluripotent cells from mouse embryos. Nature 1981; 292:154-156.; 2. Thomson J A, Itskovitz-Eldor J, Shapiro S S et al. Embryonic stem cell lines derived from human blastocysts. Science. 1998; 282:1145-1147.; 3. E. L. Herzog, C. Li, D. S. Krause, Plasticity of marrow-derived stem cells, Blood 102:3483-3493.; 4. Petersen B E, Bowen W C, Patrene K D, Mars W M, Sullivan A K, Murase N, Boggs S S, Greenberger J S, Goff J P. 1999. Bone marrow as a potential source of hepatic oval cells. Science 284:1168-1170.].
Under appropriate conditions and stimulations, bone marrow mesenchymal stem cells can differentiate into various cell lineages, such as adipocytes, osteocytes, chondrocytes, myocytes, hepatocytes, astrocytes, or neurons [5. M. Owen, A. J. Friedenstein, Stromal stem cells: marrow-derived osteogenic precursors, Ciba Found. Symp. 136 (1988) 42-60.; 6. Y. Muguruma, M. Reyes, Y. Nakamura, T. Sato, H. Matsuzawa, H. Miyatake, A. Akatsuka, J. Itoh, T. Yahata, K. Ando, S. Kato, T. Hotta, In vivo and in vitro differentiation of myocytes from human bone marrow-derived multipotent progenitor cells, Exp. Hematol. 31:1323-1330.; 7. M. C. Galmiche, V. E. Koteliansky, J. Briere, P. Herve, P. Charbord, Stromal cells from human long-term marrow cultures are mesenchymal cells that differentiate following a vascular smooth muscle differentiation pathway, Blood 82 (1993) 66.76.]. Thus pluripotent MSCs are an attractive source in the field of tissue regeneration and engineering, particularly because the use of embryonic stem cells is limited by ethical considerations.
Another advantage of MSCs is that MSCs can be easily isolated from bone marrow. These characteristics together indicate that MSCs can be used as a powerful tool in reconstructive medicine [8. S. P. Bruder, D. J. Fink, A. I. Caplan, Mesenchymal stem cells in bone development, bone repair, and skeletal regeneration therapy, J. Cell Biochem. 56 (1994) 283-294.; 9. Barry F P, Murphy J M. Mesenchymal stem cells: clinical applications and biological characterization. Int J Biochem Cell Biol. 36:568-584.; 10. Barry F P. Biology and clinical applications of mesenchymal stem cells. Birth Defects Res Part C Embryo Today. 69:250-256.; 11. Song L, Tuan R S, Transdifferentiation potential of human mesenchymal stem cells derived from bone marrow, FASEB JOURNAL 18 (6): AP 2004.; 12. Tocci, A. and Forte, L.: Mesenchymal stem cell: use and perspectives. Hematol. J., 4, 92-96 (2003); 13. M. F. Pittenger, A. M. Mackay, S. C. Beck, R. K. Jaiswal, R. Douglas, J. D. Mosca, M. A. Moorman, D. W. Simonetti, S. Craig, D. R. Marshak, Multilineage potential of adult human mesenchymal stem cells, Science 284 (1999) 143-147.; 14. Y. Jiang, B. N. Jahagirdar, R. L. Reinhardt, R. E. Schwartz, C. D. Keene, X. R. Ortiz-Gonzalez, M. Reyes, T. Lenvik, T. Lund, M. Blackstad, J. Du, S. Aldrich, A. Lisberg, W. C. Low, D. A. Largaespada, C. M. Verfaillie, Pluripotency of mesenchymal stem cells derived from adult marrow, Nature 418 (2002) 41-49.].
Although these characteristics make MSCs very promising in reconstructive medicine, some disadvantages prevent their widespread use. Generally, there are many limitation of using MSCs to clinical applications, including the low differentiation efficiency of MSCs to adult cell lineage. Part of the reason is because of the traditional 2D culture and differentiation methods. At present MSCs research field, the conventional protocols of 2D culture and differentiation method was widely applied.
There still exist some disadvantages in application of 2D monolayer culture system, although many progresses have been achieved. Because the culture area is always limited, the produced amount of differentiated MSCs from this 2D monolayer system is extremely low. Furthermore, the fate of stem cells is regulated by their niche which includes both intrinsic and extrinsic signals [15. Watt F M, Hogan B L. O. Eden: Stem cells and their niches. Science 2000; 287:1427-1430.]. The conventional protocols of 2D culture and differentiation method yields some single cells (singlets) which have been separated from their neighbors, and thus cause the misleading controlling of stem cell differentiation pathways [16. Mueller-Klieser W. Three-dimensional cell cultures: from molecular mechanisms to clinical applications. American Journal of Physiology, 273: 1109-1123.; 17. Kunz-Shughart L A, Kreutz M & Knuechel R. Multicellular spheroids: a three-dimensional in vitro culture system to study tumor biology. International Journal of Experimental Pathology, 79: 1-23.; 18. Kunz-Shughart L A. Multicellular tumor spheroids: intermediates between monolayer culture and in vivo tumor. Cell Biology International, 23: 157-161.; 19. Bates R C, Edwards N S, Yates J D. Spheroids and cell survival. Critical Reviews in Oncology/Hematology, 36: 61-74.; 20. Layer P G, Robitzki A, Rothermel A et al. Of layers and spheres: the reaggregate approach in tissue engineering. Trends in Neurosciences, 25: 131-134.].
The low differentiation efficiency limits further application of MSCs in cell therapy, tissue engineering and other regenerative medicine aspects.